Modulation of K+ channels by arachidonic acid in T84 cells. II. Activation of a Ca(2+)-independent K+ channel.
We used single-channel recording techniques to identify and characterize a large-conductance, Ca(2+)-independent K+ channel in the colonic secretory cell line T84. In symmetric potassium gluconate, this channel had a linear current-voltage relationship with a single-channel conductance of 161 pS. Channel open probability (Po) was increased at depolarizing potentials. Partial substitution of bath K+ with Na+ indicated a permeability ratio of K+ to Na+ of 25:1. Channel Po was reduced by extracellular Ba2+. Event-duration analysis suggested a linear kinetic model for channel gating having a single open state and three closed states: C3<-->C2<-->C1<-->O. Arachidonic acid (AA) increased the Po of the channel, with an apparent stimulatory constant (Ks) of 1.39 microM. Neither channel open time (O) nor the fast closed time (C1) was affected by AA. In contrast, AA dramatically reduced mean closed time by decreasing both C3 and C2. The cis-unsaturated fatty acid linoleate increased Po also, whereas the saturated fatty acid myristate and the trans-unsaturated fatty acid elaidate did not affect Po. This channel is activated also by negative pressure applied to the pipette during inside-out recording. Thus we determined the effect of the stretch-activated channel blockers amiloride and Gd3+ on the K+ channel after activation by AA. Amiloride (2 mM) on the extracellular side reduced single-channel amplitude in a voltage-dependent manner, whereas Gd3+ (100 microM) had no effect on channel activity. Activation of this K+ channel may be important during stimulation of Cl- secretion by agonists that use AA as a second messenger (e.g., vasoactive intestinal polypeptide, adenosine) or during the volume regulatory response to cell swelling.
Devor
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Department of Cell Biology and Physiology, University of Pittsburgh, Pennsylvania 15261, USA. dd2+@pitt.edu
Frizzell
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D000584:Amiloride / Q000494:pharmacology; D016718:Arachidonic Acid / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D002463:Cell Membrane Permeability; D003106:Colon; D005682:Gadolinium / Q000494:pharmacology; D006801:Humans; D007413:Intestinal Mucosa / Q000502:physiology*; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D019787:Linoleic Acid / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D019301:Oleic Acid / Q000494:pharmacology; D009829:Oleic Acids; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism / Q000494:pharmacology; D015221:Potassium Channels / Q000187:drug effects / Q000502:physiology*; D011336:Probability; D015290:Second Messenger Systems; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology
D009829:Oleic Acids; D015221:Potassium Channels; D016718:Arachidonic Acid; D019301:Oleic Acid; C011459:elaidic acid; D000584:Amiloride; D019787:Linoleic Acid; D012964:Sodium; D005682:Gadolinium; D011188:Potassium